Erythropoietin (EPO), a kidney cytokine regulating hematopoiesis, is also produced in the brain after oxidative/nitrosative stress. Hypoxia inducible transcription factor-1 (HIF-1) upregulates EPO following hypoxic stimuli. Here we propose to study preconditioning with EPO to show that it protects neurons in models of ischemic and degenerative damage due to excitotoxins and consequent generation of free radicals, including nitric oxide (NO). We propose to show that activation of neuronal EPO receptors (EPO-Rs) prevents N-methyl-D aspartate (NMDA)- and NO-induced apoptosis by triggering cross talk between the Janus kinase-2 (Jak2) and nuclear factor KB (NF-KB) signaling pathways. EPO-R - mediated activation of Jak2 leads to phosphorylation of the inhibitor of NF-KB (IKB), subsequent nuclear translocation of the transcription factor NF-KB, and NF-KB-dependent transcription of neuroprotective genes. Transfection of cerebrocortical neurons with a dominant -interfering form of Jak2 or an IKB superrepressor blocks EPO-mediated prevention of neuronal apoptosis. Thus neuronal EPO-Rs activate a neuroprotective pathway that is distinct from previously well characterized Jak and NF-KB functions. Moreover, this EPO effect may underlie neuroprotection mediated by hypoxic-ischemic preconditioning. To test this postulate, we will examine the neuroprotective properties of EPO-related molecules in a mouse middle cerebral artery occlusion model of stroke using the intraluminal suture method. The Specific Aims are as follows: To characterize EPO- induced activation of the NF-KB pathway in neuroprotection. 2. To test whether NF-KB activation following EPO exposure in mixed neuronal-glial cerebrocortical cultures occurs primarily in neurons. 3. To investigate the possibility of a direct role of EPO-activated Jak2 in NF-KB signaling. 4. To study the treatment of stroke in a mouse model with EPO-related molecules. As a proof-of principle of the involvement f o EPO signaling, we will also test these drugs as well as the effect of increased endogenous EPO in transgenic mice expressing a truncated WPO-R that is hypersensitive to EPO-induced Jak2 signaling compared to the normal full-length receptor.